The present invention relates generally to an in vitro chamber for sustaining human tissue samples during study. More particularly, this invention concerns a portable in vitro chamber adapted for use with a microscope while maintaining viability of human organ tissue samples for at least one day.
For purposes of this invention, the terms "organ tissue samples" and "tissue cultures" are to be distinguished from one another. As used herein, an "organ tissue sample" is a multicellular system such as might be obtained from a tissue biopsy or a skin specimen and includes hair, fingernails, sweat glands, and sebaceous glands. By contrast, "tissue culture" refers to a single cell type which may be nurtured for growth studies, and typically requires a substrate on which the cells grow.
Generally speaking, the concept of an in vitro chamber merely for sustaining human tissue cultures is not unique. In vitro propagation of cells conventionally occurs in culture plates or bottles manufactured from polystyrene or glass. To culture cells, the cells may be inoculated into flasks, single culture dishes or multi-well plates. A nutrient medium is added and the cells are incubated under controlled conditions. Alternatively, cells can be grown in continuously rolling bottles, on glass or polysaccharide beads, on tissue segments, or suspended in a suitable culture medium. Other methods also exist in which deforming stresses are applied to the cells to simulate cyclic stresses encountered for example in heart and lung cells.
For example, it is known in the prior art to use an apparatus having a plurality of individual wells, each having a movable membrane as the bottom wall thereof and each having a tissue culture therein. The upper surface of the membrane may be coated with an extracellular material, such as rat tail collagen, to adapt the membrane to permit attachment of tissue cells. A suitable clear plastic cover may be provided for the wells. Mechanical stretching of the membranes in each well stimulates the tissue culture. Suitable nutrients can be supplied to the tissue cultures contained in the various wells. Apparatus of this type is described more fully in U.S. Pat. No. 4,940,853 issued to Vandenburgh on Jul. 10, 1990, and U.S. Pat. No, 5,153,136 issued to Vandenburgh on Oct. 6, 1992. The present invention, however, is principally concerned with non-cyclically stressed organ tissue samples.
Other multiwell tissue culture assemblies are also known. For example, it is known to provide a plate having a multiplicity of frustoconical wells therein, each well adapted to receive a liquid cell culture medium for cell cultures. A specially designed cover may cooperate with the well plate to minimize airflow between the wells and the external environment so as to minimize evaporation, while permitting such an airflow exchange to exist thereby maintaining equilibration. U.S. Pat. No. 4,657,867 issued to Guhl et al. on Apr. 14, 1987, is an example of such an apparatus.
Multiwell apparatus have also been proposed which overcome the "edge effect" associated with some tissue culture holders by specially designing a base and cover arrangement from a suitable transparent plastic material. The base includes a plurality of tissue culture wells. A peripheral baffle around the base extends above the culture wells and is enclosed by the cover arrangement Gas may be distributed above the culture wells. To reduce uneven evaporation in the wells from air circulating around the assembly, the bottom of the base includes an insulating chamber. That insulating chamber may be filled with a clear liquid that may be either heated or cooled while permitting observation of the tissue wells by a conventional inverted microscope. Examples of these devices can be found in the Rothenberg et al. U.S. Pat. Nos. 4,673,651 issued Jun. 16, 1987, and 4,786,601 issued Nov. 22, 1988.
Various materials are also known for use in in vitro cell culture studies. As a specific example, surface-modified polyorganosiloxane compositions are known which demonstrate improved biocompatibility in cell culture apparatuses. See for example, U.S. Pat. No. 4,822,741, issued to Banes on Apr. 18, 1989.
Moreover, it is known to provide specially adapted microscope stages for optical examination of tissue culture studies. One such microscope stage having a single chamber arranged for circulation of a fluid under thermally controlled conditions. See for example U.S. Pat. No. 4,974,952 issued to Focht on Dec. 4, 1990. Another single chamber device for microscope stages also has a cylindrical chamber but provides for fluid flow through the chamber as well as thermally controlled fluid adjacent to the chamber. This device also appears to be adapted for tissue culture studies through its several references to petri dishes. See for example U.S. Pat. No. 4,195,131 issued to Papas on Mar. 25, 1980.
Automatic cultivating apparatus for tissue cultures in combination with an optical inspection assembly having a light source and various lenses is also known. Tissue culture wells are carried by an annular disk into successive registry with the optical inspection assembly. The atmosphere within the apparatus is controlled. See for example U.S. Pat. No. 4,090,921 issued Sawamura et al. on May 23, 1978.
In other technologies, microscope state assemblies have also been designed to accommodate variant temperature and pressure ranges in a single chamber sealed envelope with optical windows. See for example U.S. Pat. No. 4,707,086 issued to Dahan et al. on Nov. 17, 1987.
One device is known for maintaining single organ tissue sample where the sample rests on a concave plate with a central opening. The concave plate, in turn, is supported above a peripherally heated reservoir containing a suitable medium, a gas probe, a temperature monitor, and a magnetic stirring device. That device, however, was cumbersome, had so much peripheral matter that it could not be used on a microscope, and created a potential health hazard when operating due to splashing of medium, and exposure of the medium when the concave plate was removed.
Despite the existence of a device for maintaining in vitro a single organ tissue sample, there continues to be a need for devices which can be operatively connected with a conventional stereo microscope for examination and study. Known devices cannot be so used since associated heater assemblies interfere with placement of the container on a microscope stage. In a similar vein, complicated studies of human organ tissue require that multiple specimens be simultaneously observed and examined over periods of time. Furthermore, considering the heightened concern of exposure to HIV viruses when working with human tissue samples, there is a need to contain both the tissue samples and all fluids coming into contact with them. Recognizing that it can be desirable to begin in vitro support for organ tissue samples immediately following removal from the host body, there is also a need for a portable chamber.
Thus, a need continues to exist for an in vitro chamber adapted for use in microscope studies of human organ tissue.